The enzyme, carbonic anhydrase (“CA”) (EC 4.2.1.1), catalyzes the reversible reactions depicted in Scheme 1:

In the forward or “hydration” reaction, CA combines carbon dioxide and water to provide bicarbonate and a proton, or depending on the pH, to provide carbonate (CO3−2) and two protons. In the reverse, or “dehydration” reaction, CA combines bicarbonate and a proton to provide carbon dioxide and water. Carbonic anhydrases are metalloenzymes that typically have Zn+2 in the active site. However carbonic anhydrases having e.g. Co+2 or Cd+2 in the active site have been reported. At least three classes of carbonic anhydrases have been identified in nature.
The α-class carbonic anhydrases are found in vertebrates, bacteria, algae, and the cytoplasm of green plants. Vertebrate α-class carbonic anhydrases are among the fastest enzymes known, exhibiting a turnover number (kcat) (the number of molecules of substrate converted by an enzyme to product per catalytic site per unit of time) of 106 sec−1. The β-class carbonic anhydrases are found in bacteria, algae, and chloroplasts, while γ-class carbonic anhydrases are found in Archaea and some bacteria. Although carbonic anhydrases of each of these classes have similar active sites, they do not exhibit significant overall amino acid sequence homology and they are structurally distinguishable from one another. Hence, these three classes of carbonic anhydrase provide an example of convergent evolution.
It has been proposed to use carbonic anhydrase as a biological catalyst to accelerate the capture of carbon dioxide produced by combustion of fossil fuels. See e.g., U.S. Pat. Nos. 6,143,556, 6,524,843 B2, 7,176,017 B2, 7,596,952 B2, 7,579,185 B2, 7,740,689 B2, 7,132,090 B2; U.S. Pat. Publ. Nos. 2009/0155889A1, 2010/0086983A1; PCT Publ. Nos. WO2006/089423A1, WO2010/014773A1, WO2010/045689A1. Naturally occurring carbonic anhydrases, however, are not well-suited for use under the process relevant conditions that are required for an economically viable carbon dioxide capture system. These process relevant conditions include: presence in solution with high concentrations of other CO2 absorption mediating compounds (e.g., amines, ammonia, carbonate ions, amino acids); elevated temperatures (e.g., 40° C. or above, or 15° C. or below in NH3), alkaline pHs (e.g., pH 8-12); presence of gas contaminants (e.g., high levels NOx and SOx); and extended periods of exposure to these challenging conditions (e.g., days to weeks). In addition, such carbonic anhydrases should also be stable to variations in these process conditions, e.g., stable not only at a relatively alkaline pH suitable for hydration and sequestration of carbon dioxide but also at a relatively acidic pH suitable for subsequent release and/or recapture of the hydrated and/or sequestered carbon dioxide.
Chemical conjugates of α-class carbonic anhydrases and some of their physical properties have been described in the following references: Epton et al. “Soluble polymer-protein conjugates: 1. Reactive N-(sym-trinitroaryl)polyacrylamide/acrylhydrazide copolymers and derived carbonic anhydrase conjugates,” Polymer 18: 319-323 (1977); Farmer et al., “Assessing the Multimeric States of Proteins: Studies Using Laser Desorption Mass Spectrometry,” Biol. Mass Spectrometry 20, 796-800 (1991); Gitlin et al., “Peracetylated Bovine Carbonic Anhydrase (BCA-Ac18) Is Kinetically More Stable than Native BCA to Sodium Dodecyl Sulfate,” J. Phys. Chem. B. 110: 2372-2377 (2006); Gudiksen et al., “Eliminating Positively Charged Lysine e-NH3+ Groups on the Surface of Carbonic Anhydrase Has No Significant Influence on Its Folding from Sodium Dodecyl Sulfate,” J. Am. Chem. Soc. 127: 4707-4714 (2005); Gudiksen et al., “Increasing the Net Charge and Decreasing the Hydrophobicity of Bovine Carbonic Anhydrase Decreases the Rate of Denaturation with Sodium Dodecyl Sulfate,” Biophys. J. 91: 298-310 (2006); Bootorabi et al., “Modification of carbonic anhydrase II with acetaldehyde, the first metabolite of ethanol, leads to decreased enzyme activity,” BMC Biochemistry 9: 32 (2008); Trachtenberg et al., “Carbon Dioxide Transport By Proteic And Facilitated Transport Membranes,” Life Support & Biosphere Science 6: 293-302 (1999); and Bhattacharya et al., “CO2 hydration by immobilized carbonic anhydrase,” Biotechnol. Appl. Biochem. 38: 111-117 (2003).
Accordingly, there is a need in the art for engineered and/or chemically modified carbonic anhydrases with further improved enzymatic properties that can effectively accelerate the absorption of carbon dioxide from a gas stream and/or accelerate desorption of carbon dioxide from a capture solution under process relevant conditions.